Black-fly larvae, and mosquito larvae and pupae, are shown to select certain temperature ranges when given a choice in a linear gradient. Larval age has little effect on the temperature selected, but pupae are tolerant of higher temperatures than larvae. The significance of this observation is discussed. The temperature selected is affected by the previous thermal history of the insect, although acclimation to the ambient temperature may be rapid. The final selected temperature of fourth-instar Aedes aegypti larvae was determined. Rearing temperature has a profound effect on the thermal resistance of Aedes aegypti exposed to high temperatures. / Thesis / Master of Science (MS)

Average global ocean temperatures and the frequency and intensity of marine heat waves have been increasing over the last century. Temperature plays a critical role in defining the geographical range of the majority of marine species. Some species may respond to ocean warming trends by shifting their latitudinal and depth ranges, while others may be able to cope with changes in temperature through phenotypic plasticity and local adaptations. If a species is unable to shift its distribution or has limited thermal plasticity, it may face severe population declines or local extinction. Therefore, describing thermal tolerance limits is a useful tool for predicting how a given species will respond to ocean warming. Due to its commercial importance, sablefish (Anoplopoma fimbria) is a fish species of particular interest in British Columbia. Sablefish are semi-demersal and spawn along the continental slopes of the Pacific coast from California to Alaska. Their various life history stages occupy different depth strata and thus experience substantially different environments with respect to temperature (as well as salinity, oxygen, etc.). Adult sablefish spawn at depths that exceed 300 m and embryos sink to depths of ~1,000 m after fertilization. Embryos hatch into yolk-sac larvae until they become mobile at the post-yolk-sac larvae stage. The latter migrate to near-surface waters (<3 m) at which temperatures are approximately 12-15°C in the late spring. Heart rate is a temperature-dependent performance measure and has been used to gain insight into the thermal tolerance of many adult fishes. However, few studies have used this approach with the early life stages of fishes such as embryos and yolk-sac larvae (YSL). The purpose of this study was to assess whether sablefish embryos and YSL have the potential for cardiac acclimation by examining changes in their thermal tolerance limits when incubated at temperatures outside of what they experience in a natural setting (~5°C). Cardiac performance was assessed during an acute temperature challenge from 2.0° to 12.0°C in increments of 1.0°C (at a rate of 1°C 40 min-1) for individuals incubated at 3.0°C, 5.0°C (control), and 7.0°C. Embryos were video recorded at each 1.0°C increment and videos were viewed at a later date to determine heart rate at each temperature. This study attempted to use temperature breakpoint analysis, commonly used in studies of adults, on these early life stages to assess cardiac performance. It was hypothesized that sablefish embryos and yolk-sac larvae incubated at warmer temperatures would have a higher thermal tolerance than sablefish embryos and YSL incubated at colder temperatures, as seen in other fish species. There was some degree of thermal compensation of cardiac function with temperature in sablefish embryos and YSL as mean heart rate increased with incubation temperature throughout acute warming. YSL had consistently higher mean heart rate values at any given temperature of the acute temperature challenge when compared to embryos incubated at the same temperature. TAR is the temperature at which the heart first becomes arrhythmic is considered a sub-lethal index because the organism is expected to experience cardiac collapse soon after. TAR was reached for 100% of embryos incubated at 3.0°C at an average temperature of 8.6 ± 1.0°C. In contrast, only 18% and 33% of embryos incubated at 5.0° and 7.0°C exhibited arrhythmia (mean TAR were 9.0 ± 3.0 and 8.5± 1.5°C, respectively). The lower thermal limit for embryos incubated at 7.0°C was likely near 1.0°C, which was determined during preliminary testing. Neither the upper or lower limits were reached for YSL during the acute temperature challenge. No mortalities were observed during any acute temperature challenges. Overall percent mortality throughout the entirety of the experiment could not be determined due to limitations in the experimental setup and reduced staff working on this project due to COVID-19 safety protocols. This study is also the first to investigate whether transporting sablefish embryos from a hatchery to a research facility at different stages of development had an effect on their cardiac performance during acute warming. Embryos were transported in a cooler from the sablefish hatchery on Salt Spring Island to UVic via ferry and vehicular transport. Time of transportation did not significantly change the temperature at which heart rate reaches its maximum or TAR. There was also consistent overlap in mean heart rate ± standard error at each temperature of the acute temperature challenge between these two groups. Therefore, there appears to be no indication that transportation affected the heart rate response of sablefish embryos when incubated at the same temperature. However, future studies may want to confirm this by identifying and comparing other breakpoint temperatures that characterize physiological performance. Determining whether transportation has effects on cardiac performance may be of interest to other researchers who need to transport fish embryos from the field to the laboratory. Heart rate measurements during an acute temperature challenge of sablefish embryos and yolk-sac larvae (YSL) incubated at various temperatures provided initial insight to their overall success in a warming climate. Currently, it is projected that waters at depths of 1,000 m will warm on average by less than a degree by the end of the 21st century. The results of this study suggest that the early life stages of sablefish may not be exposed to critical temperatures in the near future, but future impacts on overall physiological decline remain unknown. The novel data presented here lay the groundwork for future researchers to continue to characterize the thermal tolerances of the early life stages of sablefish, and the likely response of this important species to ocean warming. / Graduate

The ability to acclimate to changing temperature has consequences for the biogeographic range of a species and their potential for surviving ocean warming. Using a tide simulator, which recreates tidal conditions by controlling water levels, water and air temperatures, light levels, and food availability, we explored how temperature and tidal zone (i.e. intertidal and subtidal) influences the abundance of sirtuin 5 (SIRT5) protein in a California native mussel (Mytilus californianus). We compared how gill tissue abundance of SIRT5, a key regulator of the cellular stress response and energy metabolism responded in M. californianus exposed to different temperature conditions (13, 16, 19 and 22°C) over a period of four weeks. Two SIRT5 isoforms, one a putative cytosolic form and the other a mitochondrial form were found to be expressed in mussel gill tissue. The mitochondrial isoform increased during acclimation to warm temperatures. This finding is the first to show how SIRT5 protein abundance changes with temperature acclimation. Surprisingly, we did not identify any differences in gill SIRT5 abundance between mussels from intertidal and subtidal locations. Our results suggest that characterizing the responses of SIRT isoforms may lead to a better understanding of the physiological diversity of sirtuins.

The response of global soil respiration (Rs) to climate change determines how long the land can continue acting as a carbon sink in the future. This dissertation research identifies how temporal and spatial variation in environmental factors affects global scale Rs modeling and predictions of future Rs under global warming. Chapter 1 describes the recommend time range for measuring Rs across differing climates, biomes, and seasons and found that the best time for measuring the daily mean Rs is 10:00 am in almost all climates and biomes. Chapter 2 describes commonly used surrogates in Rs modeling and shows that air temperature and soil temperature are highly correlated and that they explain similar amounts of Rs variation; however, average monthly precipitation between 1961 and 2014, rather than monthly precipitation for a specific year, is a better predictor in global Rs modeling. Chapter 3 quantifies the uncertainty generated by four different assumptions of global Rs models. Results demonstrate that the time-scale of the data, among other sources, creates a substantial difference in global estimates, where the estimate of global annual Rs based on monthly Rs data (70.85 to 80.99 Pg C yr-1) is substantially lower than the current benchmark for land models (98 Pg C yr-1). Chapter 4 simulates future global Rs rates based on two temperature scenarios and demonstrates that temperature sensitivity of Rs will decline in warm climates where the level of global warming will reach 3°C by 2100 relative to current air temperature; however, these regional decelerations will be offset by large Rs accelerations in the boreal and polar regions. Chapter 5 compares CO2 fluxes from turfgrass and wooded areas of five parks in Blacksburg, VA and tests the ability of the Denitrification-Decomposition model to estimate soil temperature, moisture and CO2 flux across the seasons. Cumulatively, this work provides new insights into the current and future spatial and temporal heterogeneity of Rs and its relationship with environmental factors, as well as key insights in upscaling methodology that will help to constrain global Rs estimates and predict how global Rs will respond to global warming in the future. / Ph. D. / CO₂ flux emitted from global soil is the second largest carbon exchange between the land and atmosphere. Accurately estimating global soil CO₂ flux and how it responds to climate change is critical to predict terrestrial carbon stocks. The objectives of this dissertation are to evaluate how time-scale affects our ability to estimate global soil CO₂ flux. In Chapter 1, we show that the best time period for measuring daily mean soil CO₂ flux is at around 10:00 am in almost all climate regions and vegetation types. The previously recommended time range (09:00 am and 12:00 pm) reasonably captures the daily mean soil CO₂ flux. The results from Chapter 2 indicate that air temperature is a good proxy for soil temperature in modeling global soil CO₂ flux. However, monthly precipitation is a uniformly poor proxy for soil water content; instead, average monthly precipitation is a better predictor for global soil CO₂ flux modeling. Chapter 3 demonstrates that the time-scale used in parameterizing models strongly affects the prediction of global CO₂ flux. When using monthly time-scale soil CO₂ flux and air temperature data, soil CO₂ flux increases as air temperature increases at air temperatures below 27 ℃, but soil CO₂ flux begins to decrease when air temperature is over 27 ℃. However, when using annual time-scale data, this response to temperature is masked, soil CO₂ flux increases as air temperature increases in all temperature conditions. As a result, the estimate of global annual soil CO₂ flux, based on monthly soil respiration data (70.85 to 80.99 Pg C yr⁻¹ ), is lower than the estimate based on the annual soil respiration data (98 Pg C yr⁻¹ ). Chapter 4 shows that if the level of global warming maintains its current rate (3ºC by the year 2100), then the annual soil CO₂ flux will either decrease or remains the same in arid, winter-dry temperate and tropical climate regions. However, these regional decelerations were offset by large soil CO₂ flux accelerations in the boreal and polar regions. Chapter 5 shows a significant difference in CO₂ flux among the five selected parks in Blacksburg, VA. The Denitrification-Decomposition model, despite having been developed for agriculture and undeveloped lands, closely estimates soil temperature, moisture and CO₂ flux across the seasons and therefore can be used to estimate and understand CO₂ fluxes from urban ecosystems in future studies. This study highlights that the relationship between soil CO₂ fluxes and environmental factors such as air temperature and precipitation differs from region to region. The study also demonstrates that daily and monthly time-scale soil CO₂ fluxes and environmental data help constrain global soil CO₂ flux estimates and help to predict how global soil CO₂ fluxes will respond to global warming in the future.

Le réchauffement climatique a des conséquences sur la phénologie, les aires de répartition et la taille des organismes. Cependant, les processus physiologiques sous-jacents à ces phénomènes restent peu connus. Les ectothermes dont le métabolisme est positivement corrélé à la température pourraient montrer, sous réchauffement climatique, une production accrue d'espèces réactives de l'oxygène (ERO). Ces ERO ont été depuis quelques décennies définis comme l'un des mécanismes physiologiques sous-jacents aux stratégies d'histoire de vie. Dans cette thèse, nous avons exploré la physiologie oxydative de poissons dans des contextes thermiques variés et ses conséquences sur la gestion des traits d'histoire de vie. Nos approches expérimentales et de terrains menées sur Oryzias latipes, Perca fluviatilis & Rutilus rutilus, nous ont permis de montrer ces liens. Ces résultats sont à compléter, notamment sur le terrain où de nombreux effets confondants ont rendu les conclusions difficiles. / Global warming affects the phenology, range and body-sizes of species. However, the underlying physiological processes remain poorly understood. Ectothermic organisms, whose metabolism is positively correlated with temperature, would show an increase in reactive oxygen species (ROS) production under global warming. These ROS have been proposed to be one of the physiological triggers of the life history strategies. In this thesis, we explored the oxidative physiology of fish in different warming contexts and their effects on life-history traits. Our experimental and field approaches on Oryzias latipes, Perca fluviatilis & Rutilus rutilus, allowed us highlight these links. However, further studies are necessary, especially in natura where many confounding effects have made conclusions difficult.

The tropical rainforests of the Peruvian Andes are some of the most biodiverse and most vulnerable to climate warming in the world. The Andes are predicted to experience substantial increases in warming of between +2 °C to +5 °C by the end of the century, in addition to an increases in the frequency of high temperature extremes, drought and flood events. The response of these forests to climate change over the next century has global relevance, due to the high levels of endemic species present and the potential role these areas will play as refugia for lowland species. Despite this, the response of tropical montane forests (TMCFs) to climate change remains under-studied. Our current understanding of how Andean species will respond to climate change is based on studies of past compositional changes. Upslope shifts in plant communities of approximately 1.2 - 2.0 m·y-1 have been observed along elevational gradients within Central and South America over the last decade. Based on these migration rates, it has been estimated that the majority of communities will lag behind increases in temperature by 5.5 °C by the end of the century. The implications of this for populations at the trailing range edge is unclear, due to a lack of mechanistic data concerning the acclimatory limits of rainforest species. When faced with rapid warming plant species will need to rapidly adapt, acclimate or migrate in order to survive. In the case of Andean species, migration rates may not be sufficient for a species to remain within its optimal thermal niche and adaptive responses will likely be too slow to be effective, hence individuals will have to acclimate in situ to prevent a decline in performance. The acclimatory ability of species can be quantified by measuring changes in performance, leaf physiology and anatomy in response to experimental manipulations of climate, however such studies are rare within the tropics. Here we carried out a seedling transplant experiment, utilising an extensive 400-3500 m asl elevational gradient in the Peruvian Andes, to simulate climate warming and upslope migration of tree seedlings under real-world conditions. To provide context for the transplant study, natural variation in leaf anatomical traits and physiological stress were explored for twelve species belonging to lowland (LF), mid-elevation (LMF) and tropical montane cloud forest communities. Adults and seedlings from the centre and furthest-most extent of each species' elevational range were studied and compared. Seedlings of each elevational forest community were transplanted downslope and upslope of their local elevational range by the equivalent of ±2 °C and ±4 °C in mean annual temperature. The experiment followed the transplanted seedlings of eleven species over a one year period, monitoring survival, growth and physiological stress (Fv/Fm) of individuals. The acclimatory ability of a subset of these species was quantified by measuring changes in photosynthetic capacity (Vcmax and Jmax), respiratory capacity (Rd) and anatomical traits (Na, Pa, LMA, LDMC) in response to transplantation. The results showed that within the natural population there was little evidence of leaf trait acclimation to elevational shifts in climate, but also little evidence of physiological stress at the trailing range edge. There were however differences in the leaf trait strategies employed by each elevational community, increasing in abiotic stress-tolerance with elevation. Physiological stress was greatest in the seedling population and, unlike the adult population, increased slightly at the trailing edge. This indicated that seedlings were more vulnerable to warming than their adult counterparts and at mid-elevations TMCF seedlings were more vulnerable than LMF seedlings. Seedling survival and growth declined in response to transplantation away from the home elevation for the majority of species, with upslope declines as a result of abiotic limitations, and downslope declines due to biotic limitations. All seedlings were found to be able to acclimate their respiratory capacity in response to transplantation, however this was not the case for photosynthetic capacity. LMF species performed significantly better than TMCF seedlings with transplantation, demonstrating a greater acclimatory capacity for photosynthesis. LMF species were able to adjust Jmax in order to maintain rates at ambient temperatures, but were not able to upregulate Vcmax upslope, whereas TMCF species were not able to respond in either transplant direction. Overall, these findings suggest that under moderate warming scenarios LMF species will have a competitive advantage over TMCF species at mid-elevations, gradually expanding their range into TMCF species' habitat over the next century. As a result of this and due to the slow pace of upslope migration, we predict that TMCF species will undergo range retractions and possible extinctions. The speed of this response will be determined by the trajectory of future warming and the frequency of extreme climatic events.

Made available in DSpace on 2014-06-11T19:30:58Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-02-26Bitstream added on 2014-06-13T20:01:15Z : No. of bitstreams: 1 portes_mt_dr_rcla.pdf: 2079689 bytes, checksum: 765fd4860ab97b37290c8d8d62e1005f (MD5) / As restrições impostas pelo ambiente obrigam as plantas a transitarem entre estados fisiologicamente distintos, podendo tal transição ser representada pelo processo de aclimatação. O aparato fotossintético apresenta alta sensibilidade ao ambiente, mas possui alta capacidade de aclimatação, a qual é necessária dada a sua função essencial no metabolismo vegetal e o seu elevado nível de integração com outras vias metabólicas. A investigação do papel da aclimatação fotossintética na resposta a diversas condições ambientais, em diferentes espécies arbóreas, foi o principal tema de estudo da presente tese. Foram realizados experimentos com espécies arbóreas nativas e cultivadas sob diferentes níveis de controle ambiental, ou seja, no campo, em casa de vegetação e em câmaras de crescimento. Os experimentos com espécies arbóreas nativas envolveram a avaliação da capacidade de aclimatação das espécies de diferentes grupos sucessionais, as quais supostamente apresentam demandas luminosas distintas e diferem em sua habilidade de ajuste e acoplamento ao ambiente. O experimento com a espécie cultivada foi realizado com Eucalyptus globulus, e avaliou o efeito da deficiência hídrica em plantas sob diferentes regimes térmicos. A conjunção dos resultados obtidos nos quatro experimentos realizados permitiu verificar que a aclimatação do aparato fotossintético foi influenciada pelo ambiente de crescimento das plantas e pela estratégia ecológica das espécies, mais conservativa ou mais flexível. Além disso, foi verificado que a estratégia ecológica das espécies não está, necessariamente, relacionada ao seu grupo sucessional, conforme freqüentemente descrito na literatura. Os diversos ajustes no aparato fotossintético, descritos no presente trabalho em diferentes espécies e condições ambientais, demonstraram... / The constraints imposed by the environment compel plants to transit between distinct physiological states, represented by the acclimation process. The photosynthetic apparatus is highly sensitive to the environment, however it presents a high acclimation capacity which is necessary given its essential role in plant metabolism and high level of integration with other pathways. The investigation of the role of photosynthetic acclimation in response of different tree species to diverse environmental conditions was the main subject of the present study. Experiments with tropical tree species and cultivated species were performed under different levels of control of environmental conditions, i.e. in the field, in the greenhouse, and in growth chambers. The experiments with tropical tree species involved the evaluation of the acclimation capacity of species belonging to different ecological groups, supposedly presenting distinct light demands and ability to adjust and couple to the environment. The experiment with cultivated species was carried out with Eucalyptus globulus and photosynthetic acclimation was evaluated under water deficit, in plants under different thermal regimes. The conjunction of the results obtained in the four experiments performed suggests that the acclimation of the photosynthetic apparatus was influenced by the growth environment jointly with the ecological strategy of the species, more conservative or more flexible. Moreover, it was verified that the ecological strategy of the species is not necessarily related with its ecological group as often stated in the literature. The diverse changes in the photosynthetic apparatus described in the present study in different species and environmental conditions, demonstrated the importance and the contribution of the photosynthetic acclimation in the physiological adjustment of a plant to its current environmental condition... (Complete abstract click electronic access below)

Le réchauffement climatique constitue une problématique d'actualité très préoccupante en raison de ses effets potentiels sur la biodiversité et le secteur agricole. Mieux comprendre l'adaptation des plantes face à ce phénomène récent représente donc un intérêt majeur pour la science et la société. L'étude de populations naturelles provenant d'un gradient d'altitude permet de corréler l'impact d'un ensemble de conditions climatiques (température, humidité, radiation, etc.) à des traits phénotypiques. Ces différentes populations sont dites adaptées à leurs conditions climatiques in natura. En cultivant ces plantes dans des conditions standardisées de laboratoire (intensité lumineuse, substrat, température, arrosage, etc...), la variabilité phénotypique observée, est alors due essentiellement à la variabilité génétique intrinsèque à chaque plante, donc à son génotype. La mise en culture de ces mêmes plantes en changeant une seule variable, par exemple la température, permet de mettre en évidence un phénotype caractéristique. Ce phénotype observé peut être une réponse d'acclimatation d'un génome adapté. Le projet WallOmics vise à caractériser l'adaptation des plantes à l'altitude par l'étude de populations naturelles d'Arabidopsis thaliana provenant des Pyrénées. Les acteurs moléculaires de l'adaptation des plantes au climat sont encore mal connus mais il apparaît que la paroi des cellules végétales pourrait avoir un rôle important dans ce processus. En effet, celle-ci représente le squelette des plantes et leur confère une rigidité tout en représentant une barrière externe sensible et dynamique face aux changements environnementaux. Sa structure et sa composition peuvent être modifiées à tout moment. Il est d'ailleurs possible de dire que cette paroi végétale donne la forme générale de la plante (taille, forme, densité, etc...), son phénotype observable. Ce projet se consacrera principalement à l'étude des parois des cellules végétales. Les nouvelles technologies ont permis l'émergence des données dites "omiques", c'est-à-dire de vastes ensembles de données provenant de niveaux biologiques multiples, comme des données écologiques, de phénotypages, biochimiques, protéomiques, transcriptomiques et génomiques. L'étude et la mise en relation de ces données ont favorisé le développement d'approches globales qui visent à établir une réponse à plusieurs échelles. C'est justement par ce type d'approche non mécanistique que le projet WallOmics a contribué à établir les bases moléculaires des modifications des parois face aux changements climatiques. / Global warming is a current issue of great concern because of its potential effects on biodiversity and the agricultural sector. Better understanding the adaptation of plants to this recent phenomenon is therefore a major interest for science and society. The study of natural populations from an altitude gradient allows correlating a set of climatic conditions (temperature, humidity, radiation, etc...) with phenotypic traits. These different populations are considered as adapted to their climatic conditions in natura. By cultivating these plants under standardized laboratory conditions (light intensity, substrate, temperature, watering, etc.), the observed phenotypic variability, is essentially due to the genetic variability intrinsic to each genotype. The growth of these same plants by changing a single variable, for example temperature, makes possible to highlight a characteristic phenotype. This phenotype may be an acclimation response of a relevant genome. The WallOmics project aims at characterizing the adaptation of plants to altitude by studying natural populations of Arabidopsis thaliana from the Pyrenees. The molecular actors of the adaptation of plants are still poorly described, but it appears that the plant cell wall could play an important role in this process. Indeed, it represents the skeleton of plants and gives them rigidity while representing a dynamic and sensitive external barrier to environmental changes. Its structure and composition can be modified at any time. It is also possible to say that the plant cell wall gives the general shape of the plant (size, shape, density, etc.), that is its observable phenotype. This project will focus mainly on the study of the plant cell wall. New technologies have enabled the emergence of the so-called "omics" data, large sets of data at multiple biological levels, such as ecological, phenotypic, metabolomic, proteomic, transcriptomic and genomic data. The study and the links between these data have favoured the development of integrative approaches aimed at establishing a response at several scales. It is precisely by this type of non- mechanistic approach that the WallOmics project has contributed to establish the molecular players of plant cell wall modifications in the global warming context.

Plants have spread to almost everywhere in the world. As they disperse, they meet many different environments to which they may be able to adapt. For a plant species to adapt to a new environment, genetic variation is needed. The individuals differ from each other in their genetic composition, which often means differences in phenotypes. Those individuals that manage to reproduce will form the next generation. With different conditions in different environments, it will not be the same phenotypes that reproduce everywhere. In that way, plant species will form into a mosaic of locally adapted populations varying genetically as the species disperses. After the last ice age plants have started to disperse away from the equators. With increasing latitudes come increasing challenges to migrating plants. As plant species disperse northwards along this gradient of varying conditions individuals are selected for cold adaptive traits like flowering time and freezing tolerance, acquired by cold acclimation. In this way, genetic variation from the original populations for these traits becomes sorted out along a latitudinal cline. The aim of this thesis was to understand how selection along a latitudinal gradient has shaped natural variation in cold adaptive traits in plants dispersing northwards, and specifically, to investigate what variation can be observed in phenotypes for these traits and how these traits correlate with genetic variation in genes known to be involved in cold acclimation. In this study significant variation was found in a sample of the model plan Arabidopsis thaliana accessions in cold adaptive traits flowering time and freezing tolerance. A clear latitudinal cline in the cold adaptive traits freezing tolerance for A. thaliana was observed. Analysis of nucleotide polymorphism for the cold responsive ICE1 (inducer of CBF expression 1) transcription factor revealed a haplotype structure with two allelic clades as well as unusually high levels of synonymous polymorphism. Nucleotide polymorphism analysis for the transcription factors CBF1, CBF2 and CBF3 (C-repeat binding factors) that play a key role in regulating the expression of a group of target genes known as the “CBF regulon” showed a distinct geographical haplotype structure. One haplotype was dominant in southern accessions while in the other northern accessions overrepresented. There was a significant effect of CBF haplotype on both freezing tolerance and flowering time even after correcting for latitude. Significant differences in CBF expression levels were found between the different CBF genes as well as between different accessions. Sequence variation at CBF was shown to have a significant effect on expression levels of CBF2. No clear correlations were found between CBF gene expression and freezing tolerance or temperature sensitivity for any of the accessions used in the study. This highlights the complex relationship between sequence variation in candidate genes and gene expression, and the problems associated with unraveling the genetic basis of ecologically important traits.

Winter rye produces ice-modifying antifreeze proteins upon cold treatment. Two of these antifreeze proteins are members of the large, highly conserved, β-1,3-endoglucanase family. This project was designed to identify glucanase genes that are expressed during cold acclimation, wounding, pathogen infection, drought or treatment with the phytohormones ethylene and MeJa. Additionally, a more detailed proteomic analysis was to be carried out to evaluate the glucanase content of the apoplast of cold-acclimated (CA) winter rye. Results of 2D SDS-PAGE analysis revealed that non-acclimated whole leaf protein extracts contain at least two β-1,3-endoglucanses while CA whole leaf protein extracts contain at least three β-1,3-endoglucanses. Subsequent 2D SDS-PAGE analysis was conducted on the apoplast extracts of NA and CA winter rye plants revealed the limitations of standard 1D SDS-PAGE. The 2-dimensional gel analysis revealed that there is a minimum of 25 proteins within the apoplast of CA winter rye, including at least 5 β-1,3-endoglucanases. Genome walking was used to isolate cold-responsive glucanase genes. The five genes isolated were designated scGlu6, scGlu9, scGlu10, scGlu11 and scGlu12. The cis-element pattern within the promoter of each gene was evaluated using online databases of documented plant cis elements. As expected, all of the promoters contained elements associated with cold, biotic and abiotic stresses, light regulation, and development. The expression patterns predicted by the cis elements in each promoter were compared to the mRNA abundance produced by each gene as detected by semi-quantitative reverse transcriptase PCR. In most cases, the abundance of transcripts arising from each gene loosely corresponded to the expression pattern predicted by the cis elements the corresponding promoter. Transcripts of scGlu9, 10 and 11 were present in cold-treated tissues and are candidates for β-1,3-endoglucanases with antifreeze activity. The results presented in this thesis provide additional insight into the apoplast proteome of CA winter rye plants as well as the complexity of the signals controlling the proteins that reside there. Although there are still a number of unresolved questions, this research opens new directions for future studies in the cold acclimation process in winter rye and specifically for the contribution of β -1,3-endoglucanses.